Maintenance of healthy blood sugar is primarily dependent upon the opposing actions of the pancreatic hormones insulin, released from [unreadable]-cells, and glucagon, released from a-cells. Secretion of glucagon from a-cells plays an essential role in countering hypoglycemia. The proposed studies are directed at understanding the regulation of a-cell function required to control glucagon secretion during hypoglycemia. Recent studies have pointed to an essential role for the melanocortin aMSH in this process. Deletion of the POMC gene in mice resulted in severe impairment of the counter- regulatory response to insulin-induced hypoglycemia due to lack of glucagon secretion. This phenotype was rescued by peripheral injection of the POMC gene product, aMSH. Preliminary studies indicate that pancreatic a-cells express an aMSH receptor, melanocortin receptor-4 (MC4R), and that a-cells display functional response to aMSH including change in intracellular [Ca2+] and glucagon secretion. The proposed study will test the hypothesis that MC4R plays a key role in the control of glucagon secretion by regulating [Ca2+]i and exocytic activity of a-cells. The studies are directed at testing this hypothesis in cultured a-cells and islets in vitro and mouse models in vivo. The studies are organized into 2 major Specific Aims. 1. Test the hypothesis that MC4R controls a-cell signaling and glucagon secretion in vitro: cAMP signaling, [Ca2+]i dynamics and glucagon secretion in cultured a-cells &islets. 2. Test the hypothesis that MC4R expressed in pancreatic a-cells is required for the counter regulatory response to hypoglycemia in vivo. This will be conducted with MC4R null mice and mice expressing MC4R only in the pancreatic a-cell. Successful completion of the studies will determine the ability of a-MSH-MC4R to control glucagon secretion and will also address potential interaction of aMSH derived signals with other cellular factors (e.g. direct glucose regulation, paracrine factors) implicated in control of glucagon secretion. This analysis of a newly described mechanism for control of a-cell secretion may point the way toward a therapeutic target for control of insulin-induced hypoglycemia and maintenance of euglycemia. This project will also allow for outstanding biomedical research experience for both undergraduates and MS students in line with the AREA-grant program objectives. PUBLIC HEALTH RELEVANCE: Hypoglycemia can be a serious complication of diabetes and can represent an impediment to intensive insulin therapy. The body's normal response to hypoglycemia is critically dependent upon the release of the hormone glucagon from pancreatic alpha cells, yet the mechanisms of alpha cell control are not well understood. The proposed studies seek to identify a major point of control of alpha cell function.